Project summary: Stem cells continuously replenish tissues and organs over an organism's lifetime through their special capacities for self-renewal and differentiation. Their behavior changes dramatically during development in response to changes in tissue size and function. For example, hematopoietic stem cells (HSCs) reside in the fetal liver prenatally. During this period, they actively proliferate and preferentially differentiate into lymphoid lineages. In later developmental stages, they migrate to the bone marrow, become mostly quiescent, and switch to a balanced differentiation program. Understanding the developmental transition of HSCs from the fetal to the adult program will provide a unique window into the regulatory mechanisms underlying tissue development and regeneration. A number of recent studies suggest that individual HSCs exhibit extensive heterogeneity in blood production during and post development. This heterogeneity plays important roles in blood production, aging and disease progression. It also makes assaying HSC development using conventional techniques difficult. To overcome the technical challenges, the proposed research will use an innovative in vivo clonal tracking technology to determine how the diverse differentiation programs of HSCs develop during the fetal-to-adult transition at the clonal level. We also determine the molecular mechanisms underlying the development of individually diverse HSC clones. The proposed research will investigate HSC development from a unique clonal perspective. This innovative approach will allow us to identify HSC regulatory factors that are undetectable at the population level and to discover new strategies for controlling HSC differentiation. This study will also provide a new perspective for understanding and treating hematological diseases associated with specific developmental stages. The ultimate goal of the proposed research is to harness the mechanisms that underlie the differential development of individual stem cells to improve tissue regeneration and to advance precision medicine in stem cell-related therapies.